The goal of this project is the development and evaluation of new, solution-state NMR methodologies for use in studies of structure and dynamics of biomolecules. NMR spectroscopy is an extremely powerful tool for the study of biomolecules in solution, due to its ability to provide detailed information at an atomic level. The extent to which NMR spectroscopy can be exploited for the structure determination of biomolecules, the investigation of intermolecular interactions in systems such as drug/receptor, enzyme/substrate and antibody/antigen complexes, and the characterization of biomolecular dynamics, depends on the ability to manipulate the relevant NMR-active nuclei to yield the desired information. The enormous impact that NMR spectroscopy has made during the last decade in the investigation of biomolecules and biophysical processes is due largely to the continuing development of new or improved techniques for extracting useful data from the systems of interest. The proposal targets severe general areas of solution-state NMR spectroscopy in which a number of specific goals will be pursued: 1) Development of coherence transfer experiments that utilize dipolar couplings will be explored. With the recent introduction of methods for effecting slight anisotropies in molecular tumbling, there is a wealth of new structural information potentially available from the exploitation of the residual couplings that are generated in these partially aligned samples. 2) Improvements in TOCSY experiments via the development of optimized, adiabatic mixing sequences will be investigated. This work is of significant interest in ultra-high field NMR applications due to the possibility of improving coherence transfer -bandwidths while minimizing RF sample-heating. 3) The use of cryogenic probe technology will be examined in applications that focus on obtaining valuable structural information in conformationally flexible regions of a molecule or complex, where chemical exchange broadening normally impedes conventional measurements. 4) A new technique for investigating chemical and conformational exchange effects will be applied to the study of dynamical processes in an RNA hairpin. Understanding how RNA molecules function will require detailed knowledge of their internal dynamics. The possibility for further developments of this technology, such that it could be applied to fully 13C-labeled molecules, will be explored. 5) Our existing simulation software will be improved to allow a determination,of nuclear spin dynamics under the influence of RF irradiation, spin relaxation and exchange processes. Such theoretical tools are essential for developing the experimental techniques described in this proposa1 and for correctly interpreting much of the data that will be recorded. The correct interpretation of NMR data depends critically on having a detailed understanding of the underlying spin physics of the experiments.